Study On The Determination Of Residual Contamination Of Estrogens In Animal Derived Food And Water

Nowadays, the direct or indirect influences of estrogenic chemicals on the environment has been considered as an increasingly severe problem, which threaten the health of Chinese households and the nation's foreign trade, although the abuse of estrogenic chemicals in food producing animals has been banned pretty long time ago. But up to now, the national standard of examination method for estrogens has not been published yet, except diethylstilbestrol. So it is necessary to establish a set of complete and accurate methods to control and determinate the estrogen residues in animal derived food and water.In the present study, a high performance liquid chromatographic method has been established for the determination of five estrogens (estradiol, estriol, estrone, ethynylestradiol, and diethylstilbestrol) in meat of livestock, poultry and water. The meat sample was extracted with Tris and methanol, the extraction was purified and concentrated by Oasis HLB solid phase extraction column (6 ml/200 mg). The water sample was purified and concentrated by Alhech Extract-clean columns C18 solid phase extraction column(200mg/4.0ml). They are detected and determined by reversed-phase high-performance liquid chromatography and the chromatographic conditions are as follows: Eclipse XDB-Cl8 column (5μm,4.6mm×150mm) with the temperature of 25℃,using (A)acetonitrile-(B)water as mobile phase with the flow rate of 1mL/min,an initial hold at 35% A for 3min,then slowly raised to 52% A during the following 4 min, and hold 52% A for 8min. The UV wavelength used for detection was set at 210 nm. Good separation was achieved in the 5 kinds of estrogens and the calibration curve showing a direct ratio between the concentration and the peak area were obtained,r>0.9993. Method detection limits were in the range of 14～18ng/L(water) and 3.9～4.9ng/g(meat). The method precision and accuracy were satisfactory with recovery percentages ranging from 68% to 106% and the relative standard deviations lower than 6%.Detecting different samples such as pork, chicken, underground water and surface water by above method. The results showed that four estrogens (estriol, estrone, ethynylestradiol, and diethylstilbestrol) were detected. The positive samples were confirmed by LC-MS/MS , by far the best technique for obtaining structural information about estrogens. And the fragmentation pathway and proposed fragmentation pattern for 5 kinds of estrogens were studied preliminarily. The confirmatory test provided reliable basis for the establishment of experimental method in this article. It shows that this method is accurate, rapid and simple, suitable for trace analysis in animal derived food and water.